• Journal of Ginseng Research
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• v.35 no.3
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• pp.272-282
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• 2011

Ginseng (Panax ginseng Meyer) has been shown to have anti-aging effects in animal and clinical studies. However, the molecular mechanisms by which ginseng exerts these effects remain unknown. Here, the anti-aging effect of Korean red ginseng (KRG) in rat testes was examined by system biology analysis. KRG water extract prepared in feed pellets was administered orally into 12 month old rats for 4 months, and gene expression in testes was determined by microarray analysis. Microarray analysis identified 33 genes that significantly changed. Compared to the 2 month old young rats, 13 genes (Rps9, Cyp11a1, RT1-A2, LOC365778, Sv2b, RGD1565959, RGD1304748, etc.) were up-regulated and 20 genes (RT1-Db1, Cldn5, Svs5, Degs1, Vdac3, Hbb, LOC684355, Svs5, Tmem97, Orai1, Insl3, LOC497959, etc.) were down-regulated by KRG in the older rats. Ingenuity Pathway Analysis of untreated aged rats versus aged rats treated with KRG showed that the affected most was Cyp11a1, responsible for C21-steroid hormone metabolism, and the top molecular and cellular functions are organ morphology and reproductive system development and function. When genes in young rat were compared with those in the aged rat, sperm capacitation related genes were down-regulated in the old rat. However, when genes in the old rat were compared with those in the old rat treated with KRG, KRG treatment up-regulated C21-steroid hormone metabolism. Taken together, Cyp11a1 expression is decreased in the aged rat, however, it is up-regulated by KRG suggesting that KRG seems enhance testes function via Cyp11a1.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.32 no.3
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• pp.345-352
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• 1999

The sand dollar, Astriclypeus manni is commonly encountered on a subtidal sand bottom along the coast of Cheju Island. limited information has been reported on their ecology despite their natural abundance. This study reports ecology and an annual reproductive cycle of the sand dollars present at Hamdock, northern coast of Cheju Island. A. manni collected from Hamdock revealed that they are 80 to 200 mm in test diameter. Sediment Brain size analysis indicated that A. manni mostly occurs on medium (particle diameter of 500 $\mu$m) to very find sand (particle diameter of 125 $\mu$m), particularly on fine sand (particle diameter of 250 $\mu$m). Internal morphology and in situ observations on their feeding habit indicated that A. manni is a deposit feeder, feeding on organic debris contained in the sediment around its habitat. A. manni were more frequently observed near Zostrea marina bed where content of organic matter in the sediment is considered to be higher. Gonadal tissues of the male were yellow in color while female gonads appeared to be purple. Fully mature eggs, with a mean diameter of 381 $\mu$m, and sperm were observed from the histological slides of the sand dollars collected in late July to August, suggesting that A. manni spawn during July to August when water temperature reaches 20 to $25^{\circ}C$.

• Development and Reproduction
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• v.2 no.1
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• pp.89-99
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• 1998

This study was carried out to investigate the morphological changes of gonadotropes in pituitary gland and spermatogenic cells in testis, obtained from 150 of 3-year-old immature and mature male rainbow trout (Oncorhynchus mykiss) during the reproductive cycles from March to February in the following year. In the maturation cycle of the pituitary gonadotropes of cultured rainbow trout, three periods can be distinguished i.e. a period of resting(March-August), a period of full spermatogenesis (September-November), and a period of breeding (December-February). The ultrastructures of the gonadotropes largely parallel the cyclical changes in the tests. The seminiferous tubules contain all spermatogenetic stages and sperm cells in a period of early maturation. At first, the size of the nucleus and cytoplasm decrease gradually at every stages from spermatogonia to spermatids. In the secondary spermatocytes, the small mitochondria are located over the outer cytoplam. In spermatids, the cytoplasmic masses move toward the posterior part of the nucleus. In spermatids, the two large mitochondria are located over the cytoplasm. In spermatids, the cytoplasmic masses move towark the posterior part of the nucleus. In spermatids, the two large mitochondria are located over the cytoplasm and begin to elongate. In spermatozoa, the surface of the nucleus devreases in volume. Examination by TEM shows that the nuclear envelope and plasma membrane are slightlywrinkled and closely adhered to the nucleus of spermatozoa. Two oval mitochondria are quite separated and the flagellum is inserted into the base of the spermatozoa head.The axoneme in this fish has the typical pattern such as nine peripheral doublets and a central doublet(9+2). there are remarkable individual differences in the size and morphology of spermatozoa head as observed by transmission and scanning electron microscopy.

• Reproductive and Developmental Biology
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• v.35 no.2
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• pp.137-141
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• 2011

These study was to investigate the in vitro fertilization and viability of fresh and vitrified oocytes. Also, the developmental capacity of IVF and intracytoplasmic sperm injection (ICSI) oocytes were investigated. Then vitrification was performed with the use of 20% ethylene glycol + 20% DMSO + 0.5 M sucrose + 10% FCS + TCM-199 medium. Vitrification immature oocytes are cultured in vitrification solution for 10 min afterwards transferred to expose at room temperature for 5 min. and transferred to the ice water for 5 min. The oocytes were sealed in a 1.0 mm straw and placed in a $LN_2$ container. Frozen oocytes were rapidly thawed in a water bath at $30{\sim}35^{\circ}C$, and then placed in TCM-199 medium containing 0.5 M sucrose for 5 min each, respectively, at $38^{\circ}C$. After being washed for 2~3 times, using fresh medium the oocytes were cultured in TCM-l99 medium supplemented with 5% FCS at $38^{\circ}C$ in 5% $CO_2$ and air. The normal morphology of fresh and vitrified-thawed oocytes were $87.1{\pm}2.1%$ and $54.8{\pm}2.5%$, respectively. The viability rates of fresh and vitrified-thawed oocytes were $70.0{\pm}2.2%$ and $41.9{\pm}2.6%$, respectively. Viability rates of vitrified-thawed oocytes were lower than that of fresh follicular oocytes (p<0.05). The in vitro maturation rates of fresh and vitrified oocytes were $45.1{\pm}3.6%$ and $28.9{\pm}4.4%$, respectively. The IVF rates of fresh follicular and vitrified-thawed oocytes were 34.00.2% and $20.2{\pm}2.6%$, respectively. The in vitro maturation and fertilization rates of vitrified-thawed oocytes were lower than those of the fresh follicular oocytes (p<0.05). A total of 350 oocytes were fixed and stained after co-incubation with spermatozoa, of which 88 had identifiable nuclear material. After IVF for 20 hrs, $25.1{\pm}3.4%$ of the oocytes found to have been penetrated by spermatozoas. Oocytes were fixed and stained after ICSI, and 105 oocytes contained identifiable nuclear material. After IVF and ICSI for 20 hrs, $34.3{\pm}3.4%$ and $59.0{\pm}2.0%$ of the oocytes were found to have been penetrated by spermatozoas. The developmental rates upon ICSI were significantly higher than those of the IVF method (p<0.05).

• Development and Reproduction
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• v.3 no.1
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• pp.53-58
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• 1999

The androgenic gland secretes a hormone, androgenic gland hormone, which is believed to act on the differentiation of the primary, secondary, and behavioral sex characteristics in most malacostracan crustaceans. This report presents the ultrastructural morphology of the androgenic gland in the freshwater prawn Macrobrachium nipponense. This gland, located in the coxopodite of the last pair of walking legs, was attached to the subterminal region of the sperm duct. The gland was composed of simple cellular strands, encased by a fibrous sheath. Microvilli were situated in the fibrous sheath, especially at the edge of each cellular strand. The androgenic gland cells had the large and round nucleus and rough endoplasmic reticulum arranged either in spirals or in concentric circles throughout the cytoplasm of the cell. They also had the well-developed Golgi complex and long mitochondria with flat and transverse cristae. The Golgi complex was similar to microvesicular cluster, but usually in the shape of typical dictyosomes, These features of androgenic gland cells coincides well with the protein/peptide secretion in their function. However, despite the apparent ultastructual equipment for protein/peptide secretion, no accumulation of materials secreted were noticed in the cytoplasm. Therefore it is strongly suggested that the transient transportation of the materials into the hemocoel has occurred just after synthesis.

• Applied Microscopy
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• v.41 no.1
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• pp.37-45
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• 2011

Known as a female hormone, estrogen, has an effect on the male reproductive organs. The estrogen has to combine with the estrogen receptor to communicate a signal. Propyl pyrazole triol (PPT) is an estrogen receptor alpha selective agonist with a 410-, or 1,000-fold relative binding affinity for estrogen receptor alpha versus estrogen receptor beta. In this study, adult male mice were treated weekly with subcutaneously injections of PPT (0.01 mg, 0.1 mg, 1mg and 4 mg) suspended in castor oil (as control) for 8 weeks and observed histologically changes in testis, efferent ductule and epididymis. In the high concentrations of PPT 4 mg treatment group, a remarkable reduction was observed in the weight of the body, testis and epididymis. Microscopic examination revealed a reduction in seminiferous tubular diameter of the testis, and epithelial cell height of the epididymis in treated group during the experiment. In addition, as the diameter of the efferent ductule increased gradually, the height of epithelial cells was decreased. PPT 4 mg treatment group caused inhibition of spermatogenesis due to atrophied germinal epithelium in the testis, and decrease of adipocyte size attached to the epididymis. Sperm was not observed in the caudal epididymis of PPT 4 mg treated group. In conclusion, the injection of high concentrations of PPT into adult male mice induced physiological changes, such as an inhibition of spermatogenesis, and also histological changes within the reproductive organs.

• Applied Microscopy
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• v.35 no.2
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• pp.89-96
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• 2005

The ultrastructures of the fertilized egg envelope from Hyphessobrycon serpae belonging to Characidae was studied using scanning and transmission electron microscopes to get systematic fundamental data for classification of species and to confirm whether micropyle is a common trait of Characidae or not. The fertilized egg was of colorless, transparent, spherical, adhesive and demersal type. There were not oil droplets in vitelline membrane and attached structures in the outside of fertilize egg envelope. The egg envelope had a single micropyle resembling the pathway of sperm in the area of the animal pole. The micropyle was surrounded by 13 to 15 protruded lines of the egg envelope in a radiated form. The outer surface of fertilized egg envelope was covered by reticular adhesive fibrous structures and irregularly arranged by pore canals. The fertilized egg envelope consisted of three distinct layers an outer adhesive fibrous layer with high electron density, a middle layer with pore canals, and an inner layer consisting of 6 to 7 lamellae alternating layers with interlamellae of lower electron density. These ultrastructural characters of fertilized egg envelope form Hyphessobrycon serpae can be utilized in taxonomy of teleost, and as fundamental data for study on early development of fertilized egg. It seems that the morphology of micropyle is a common trait of Characidae

• Korean Journal of Fisheries and Aquatic Sciences
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• v.33 no.5
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• pp.403-407
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• 2000

Ultrastructure and function of testis somatic cells in freshwater prawns Macrobrachium nipponense were studied. The paired testes of the prawn were elongated, united at their anterior end, which lay between the dorsal surface of the hepatopancreas and the heart. Each testis consisted of a large number of seminiferous cords compactly held together by connective tissue. A seminiferous cord was composed of an outer layer of simple squamous epithelium, a basement membrane, the closely packed germ cells and sustentacular cells of the germinal ridge, and an inner layer of simple cuboidal epithelial cells. Leydig cell-like cells in an angular areas filling the space of the seminiferous cords were observed. The nuclei of leydig cell-like cells were characterized by a distinct nucleolus. The simple squamous epithelial layer was composed of flattened cells tying on a basement membrane. The nuclei of the flattened cells were often overlapped in a layer, and the cytoplasm of the cells was observed just near the nuclei. The sustentacular cells were complex in morphology. These cells had relatively small cell bodies from which long cytoplasmic extensions ramified reached the space of germ cells in the germinal ridge. The nuclei of sustentacular cells usually exhibited angular profiles and located most commonly at the periphery of the cords. Cells of simple cuboidal epithelium located between germinal ridge and lumen of seminiferous cord, and part of the cells were adjacent to basal lamina, The cuboidal epithelial cells contained numerous mitochondria, the well-developed rER, the well-developed Golgi complex, and irregularly shaped nuclei. Transition vesicles appeared on the cis side of the Golgi complex. The large vesicles on the trans side of the complex appeared to fuse to form a membrane-bound structure. A number of pits on the cell apex suggested exocytotic activity for secretion of the sperm supporting matrix.

• Journal of Embryo Transfer
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• v.21 no.3
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• pp.255-262
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• 2006

The purpose of this study is to investigate the effects of vitrification in open pulled straws (OPS) methods on in vitro survival ability of porcine embryos. For in vitro maturation of immature oocytes, the porcine ovaries were collected from local slaughter-house. The cumulus-oocytes complexes were aspirated from 2 to 6 mm follicles. The collected oocytes were cultured for in vitro maturation in NCSU-23 medium with 5 mM hypotaurine, 0.57 mM cysteine, 10% porcine follicle fluid, 10 IU/ml PMSG and 10 IU/ml hCG for $21{\sim}22$ hrs. Then, the oocytes were more cultured $21{\sim}22$ hrs in vitro maturation in medium removed hormones. The frozen-thawed spermatozoa were washed by centrifugation 2 times for 10 min at 1,500 rpm in D-PBS with 5.56 mM glucose, 0.33 mM Na-pyruvate, 100 IU/ml penicillin, $100 {\mu}g/ml$ streptomycin and 4 mg/ml BSA. The fertilization medium used mTBM with 2 mM caffeine and 2 mg/ml BSA and adjusted to a pH of 7.2 to 7.4. The final concentration of spermatozoa was adjusted to $2.5{\times}10^6$cells/ml motile sperm during fertilization in vitro. At 8 hrs after insemination, the oocytes were transferred into NCSU-23 medium with 5.0 mM hypotaurine, 4 mg/ml BSA and 10 ng/ml EGF and cultured for 7 days. When the blastocysts of different stages were frozen-thawed by OPS methods, the proportions of embryos with normal morphology were significantly (p<0.05) higher in embryos frozen-thawed at expanded blastocyst stage (38.9%) than in early blastocyst stage (28.3%). On the other hand, the proportions of embryos damaged after frozen-thawing were significantly (p<0.05) higher in embryos frozen at early blastocyst stages than in expanded blastocyst stage. In another experiment, the normal embryos morphology after frozen- thawing were further cultured for 48 hrs. After culture, the proportions of embryos hatched were 6.7, 20.0 and 33.3% for embryos frozen-thawed at early blastocyst, mid-blastocyst and expanded blastocyst stages. These finding indicate the possible broader application for OPS methods, as frozen-thawed embryos may be accompanied by developmental stage according to requirements of the survival ability after freezing of blastocyst stage in the pig.

• Development and Reproduction
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• v.6 no.2
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• pp.97-104
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• 2002

In most mammals, mature oocyte-cumulus complexer(OCCs) ovulate into the oviduct where fertilization by sperm takes place. However, the complex that fail to fertilize eventually undergoes degeneration while they reside in the oviduct. Yet there is no blown mechanism how both oocyte and cumulus cells degenerate. Using human follicular fluid (hFF), bovine oviductal tissue extract (BOX) and mouse OCC, the present study aimed to find how the oviduct influence the viability of the oocyte and cumulus cells in vitro. There was no difference of oocyte maturation rate between the control and BOX-treated groups. However, there was a significant difference in the survival of cumulus cells between two groups. Cumulus cells cultured in the presence of hFF alone underwent initially expansion and then they formed monolayer in the culture dish. Even after 72 hr, they proliferated well and showed fibroblast-like morphology. Cumulus cells cultured in the presence of both hFF and BOX also expanded after 24 hr, however, after 72 hr culture, they eventually detached and degenerated. Cumulus cells cultured in the BOX alone gave a similar drastic result. When the cumulus cells cultured in the presence of BOX were stained with DAPI, their nuclei showed partial condensation and fragmentation. After detailed analysis of these cells by TUNEL assay, many nuclei of them exhibited well stained spots indicating the signs of apoptosis. Based upon these observations, it is suggested that BOX might possess a factor that leads mouse cumulus cells to undergo apoptosis in vitro.